IN-VITRO AND IN-VIVO CHARACTERIZATION OF NK3 RECEPTORS IN THE RABBIT EYE BY USE OF SELECTIVE NONPEPTIDE NK3 RECEPTOR ANTAGONISTS

1 Inhibition of NK3 receptor agonist-induced contraction in the rabbit isolated iris sphincter muscle was used to assess the in vitro functi onal activity of three 2-phenyl-4-quinolinecarboxamides, members of a novel class of potent and selective non-peptide NK3 receptor antagonis ts. In addition, an in vivo correlate of this in vitro response, namel y NK3 receptor agonist-induced miosis in conscious rabbits, was charac terized with some of these antagonists. 2 In vitro senktide (succinyl- [Asp(9),MePhe(8)]-substance P (6-11) and [MePhe(7)]-neurokinin B ([MeP he(7)]-NKB) were potent contractile agents in the rabbit iris sphincte r muscle but exhibited quite different profiles. Senktide produced mon ophasic log concentration-effect curves with a mean PD2 = 9.03 +/- 0.0 6 and mean n(H) = 1.2 +/- 0.02 (n = 14). In contrast, [MePhe(7)]-NKB p roduced shallow log concentration-effect curves which often appeared b iphasic (n(H) = 0.54 +/- 0.04, n = 8), preventing the accurate determi nation of pD(2) values. 3 The contractile responses to the NK3 recepto r agonist senktide were antagonized in a surmountable and concentratio n-dependent manner by SB 223412 benzyl)-3-hydroxy-2-phenylquinoline-4- carboxamide; 3-30 nM, pA(2) = 8.4, slope = 1.8 +/- 0.3, n = 4), SB 222 200 lbenzyl)-3-methyl-2-phenylquinoline-4-carboxamide; 30-300 nM, pA(2 ) = 7.9, slope = 1.4 +/- 0.06, n = 4) and SB 218795 xycarbonylbenzyl)- 2-phenylquinoline-4-carboxamide; 0.3 and 3 mu M apparent pK(B) = 7.4 /- 0.06, n = 6). 4 Contractile responses to the NK3 receptor agonist [ MePhe(7)]-NKB in the rabbit iris sphincter muscle were unaffected by S B 218795 (0.3 and 3 mu M, n = 8). In contrast, SB 223412 (30 and 300 m u M, n = 4) and SB 222200 (0.3 and 3 mu M, n = 4) inhibited responses to low concentrations (less than or equal to 1 nM), to a greater exten t than higher concentrations (>1 nM) of [MePhe(7)]-NKB. Furthermore, l og concentration-effect curves to [MePhe(7)]-NKB became steeper and mo nophasic in the presence of each antagonist. 5 SB 218795 (3 mu M, n = 4) had no effect on contractions induced by transmural nerve stimulati on (2 Hz) or substance P, exemplifying the selectivity of this class o f antagonist for functional NK3 receptors over NK1 receptors in the ra bbit. 6 In vivo, senktide (1, 10 and 25 mu g i.v., i.e. 1.2, 11.9 and 29.7 nmol, respectively) induced concentration-dependent bilateral mio sis in conscious rabbits (maximum pupillary constriction = 4.25 +/- 0. 25 mm; basal pupillary diameter 7.75 +/- 0.48 mm; n = 4). The onset of miosis was within 2-5 min of application of senktide and responses la sted up to 30 min. Responses to two i.v. administrations of 25 mu g se nktide given 30 min apart revealed no evidence of tachyphylaxis. Topic al administration of atropine (1%) to the eye enhanced pupillary respo nses to 25 mu g senktide. This was probably due to the mydriatic effec t of atropine since it significantly increased baseline pupillary diam eter from 7.0 +/- 0.4 mm to 9.0 +/- 0.7 mm (n = 4), thereby increasing the maximum capacity for miosis. Senktide-induced miosis was inhibite d by SB 222200 (1 and 2 mg kg(-1), i.v., i.e. 2.63 and 5.26 mu mol kg( -1); maximum inhibition 100%; n = 3-4), SB 223412 (0.5 and 1 mg kg(-1) , i.v., i.e. 1.31 and 2.61 mu mol kg(-1); maximum inhibition 100%; n = 3), SB 218795 (0.5 and 1 mg kg(-1), i.v., i.e. 1.26 and 2.52 mu mol k g(-1); maximum inhibition 78%; n = 3), and the structurally distinct N K3 receptor antagonist SR 142801 )4-phenylepipiperidin-4-yl)-N-methyla cetacetamide; 1.5 mg kg(-1), i.v., i.e. 2.47 mu mol kg(-1), maximum in hibition 92%; n = 3). 7 Topical administration of senktide (25 mu g; 2 9.7 nmol) to the eye induced unilateral miosis in the treated eye only . At this dose there was no significant difference (P < 0.05) between pupillary constriction obtained by topical or i.v. senktide, and topic ally administered atropine had no significant effect on responses to t opical senktide (n = 4). 8 [MePhe(7)]-NKB (125, 250 and 500 mu g, i.v. , i.e. 98.31, 196.62 and 393.24 nmol, respectively) also induced bilat eral miosis in conscious rabbits (maximum pupillary constriction = 4.1 3 +/- 0.30 mm; n = 4), but in contrast to in vitro studies this agonis t was approximately 100 fold less potent than senktide. [MePhe(7)]-NKB -induced miosis was inhibited by SB 222200 (5 mg kg(-1), i.v., i.e. 13 .14 mu mol kg(-1); maximum inhibition 69%; n = 3). 9 In summary, SB 22 3412, SB 222200 and SB 218795 are potent and selective antagonists of NK3 receptor-mediated contraction in the rabbit isolated iris sphincte r muscle. In addition, NK3 receptor agonist-induced miosis in consciou s rabbits is a good in viva correlate of the in vitro rabbit iris sphi ncter muscle preparation and appears to be a useful model for characte rizing the pharmacodynamic profile and efficacy of structurally distin ct NK3 receptor antagonists, such as SB 222200, SB 223412, SB 218795 a nd SR 142801.